LEQ-506

Time is a critical factor in drug action. The duration of inhibition of the target or residence time of the drug molecule on the target often guides drug scheduling.

The effects of time on the concentration-dependent cytotoxicity of approved and investigational agents [300 compounds] were examined in the NCI60 cell line panel in 2D at 2, 3, 7 and in 3D 11 days.

There was a moderate positive linear relationship between data from the 2-day NCI60 screen and the 3-, 7- and 11-day and a strong positive linear relationship between 3-, 7- and 11-day luminescence screen IC₅₀s by Pearson correlation analysis. Cell growth inhibition by agents selective for a specific cell cycle phase plateaued when susceptible cells were growth inhibited or killed. As time increased the depth of cell growth inhibition increased without change in the IC₅₀. DNA interactive agents had decreasing IC₅₀s with increasing exposure time. Epigenetic agents required longer exposure times; several were only cytotoxic after 11 days' exposure. For HDAC inhibitors, time had little or no effect on concentration response. There were potency differences amongst the three BET bromodomain inhibitors tested, and an exposure duration effect. The PARP inhibitors, rucaparib, niraparib, and veliparib reached IC₅₀s < 10 μM in some cell lines after 11 days.

The results suggest that variations in compound exposure time may reflect either mechanism of action or compound chemical half-life. The activity of slow-acting compounds may optimally be assessed in spheroid models that can be monitored over prolonged incubation times.

The smoothened (SMO) receptor, an essential signal transducer in the Hedgehog pathway, was targeted with antagonists to suppress the tumor. It is interesting that SMO D473H mutation confers resistance on inhibitor LDE-225 rather than LEQ-506. In this paper, the binding modes of them against the wild type and mutant SMO receptors were identified to gain insights into the resistant and non-resistant factors, based on a comprehensive protocol involving molecular docking, molecular dynamic simulations, free energy calculation and decomposition. A comparison of resistant LDE-225 and non-resistant LEQ-506 indicates that the volume of the binding cavity decreases seriously in the mutant complex with resistant LDE-225. In addition, the D473H mutation disrupts the hydrogen bond network with residues R400 and Q477, which results in the TM6 conformation inward. Owing to the absence of the hydrogen bond, residues R400 and Q477 make weak contributions to LDE-225. However, the D473H mutation along with TM6 conformational change has no effect on non-resistant LEQ-506. Finally, the resistance ascribes to adverse interaction between the greater polarity of mutant residue H473 and the nonpolar phenmethyl of LDE-225. The elaborate insights into structural and energetic mechanism of drug resistance provide an effective strategy to design rationally non-resistant antagonists.